Optical Manipulation of Gb 3 Enriched Lipid Domains: Impact of Isomerization on Gb 3 -Shiga Toxin B Interaction

The plasma membrane is a complex assembly of proteins and lipids that can self-assemble in submicroscopic domains commonly termed "lipid rafts", which are implicated in membrane signaling and trafficking. Recently, photo-sensitive lipids were introduced to study membrane domain organization, and photo-isomerization was shown to trigger the mixing and de-mixing of liquid-ordered (l ) domains in artificial phase-separated membranes. Here, we synthesized globotriaosylceramide (Gb ) glycosphingolipids that harbor an azobenzene moiety at different positions of the fatty acid to investigate light-induced membrane domain reorganization, and that serve as specific receptors for the protein Shiga toxin (STx). Using phase-separated supported lipid bilayers on mica surfaces doped with four different photo-Gb molecules, we found by fluorescence microscopy and atomic force microscopy that liquid disordered (l ) domains were formed within l domains upon trans-cis photo-isomerization. The fraction and size of these l domains were largest for Gb molecules with the azobenzene group at the end of the fatty acid. We further investigated the impact of domain reorganization on the interaction of the B-subunits of STx with the photo-Gb . Fluorescence and atomic force micrographs clearly demonstrated that STxB binds to the l phase if Gb is in the trans-configuration, whereas two STxB populations are formed if the photo-Gb is switched to the cis-configuration highlighting the idea of manipulating lipid-protein interactions with a light stimulus.